Project summary: Hydroxylamine and nitroso metabolites have been implicated in the pathogenesis of adverse reactions to sulfamethoxazole (SMX) and other arylamine drugs. Hydroxylamines spontaneously oxidize to reactive nitroso metabolites, which, in the case of SMX, can trigger delayed-type hypersensitivity reactions. These hypersensitivity reactions interfere with the effective use of SMX, which is the drug of choice for the prevention of opportunistic infections in immunocompromised patients. Studies of metabolic risk factors for SMX hypersensitivity have largely yielded negative results; however, these studies have not considered the reductive metabolism of hydroxylamine and nitroso metabolites. We have recently shown that hydroxylamines are detoxified by the flavoprotein NADH cytochrome b5 reductase (b5R) and the hemeprotein cytochrome b5 (cyt b5), through a novel, direct pathway of xenobiotic reduction. We have also shown that ascorbate, in addition to thiols, provides a major pathway of nitroso reduction. Our overall hypothesis is that impaired hydroxylamine or nitroso reduction predisposes patients to adverse reactions to arylamine compounds, such as hypersensitivity to SMX. We will address this hypothesis by first characterizing variability in hydroxylamine reduction and its relationship to genetic polymorphisms in b5R or cyt b5 in humans. We will next determine whether alterations in hydroxylamine or nitroso reduction influence the immunogenic response to SMX metabolites, using ascorbate, thiol, or flavin restriction in a guinea pig model. Finally, we will determine whether impaired hydroxylamine or nitroso reduction is a risk factor for SMX hypersensitivity, in a prospective study of immunocompromised patients with lymphoid malignancies. These studies will advance our understanding of the mechanisms underlying SMX hypersensitivity, and will characterize individual variability in a novel direct pathway of xenobiotic reduction, with clinical implications for responses to many compounds, including amidoxime pro-drugs and arylamine carcinogens, in addition to SMX. Relevance: These studies will help us to better understand individual risk factors that lead to "sulfa drug" allergies in people, by learning about differences in the ways that sulfa drugs are metabolized by different people. The ultimate goal of these studies is to find better ways to prevent these adverse reactions.